Mechanosignaling

How do focal adhesions mature?

Adhesions that manage to grow beyond the transient phase further undergo qualitative changes in a phase described as maturation. This transformation of adhesion sites into stress-fiber bound focal adhesions (FAs) is believed to result from the collective effects of Rho GTPase activity [1][2][3], increased tyrosine kinase activity [4][5] (reviewed in [6]), change in composition [7][8] and dynamics [9], actin crosslinking by α-actinin [10]and tension-dependent actomyosin contractility ([11][12][13], reviewed in [14][15]) – all of which take place during local contraction of the lamellipodium [8] (reviewed in [16]).

Rho kinase (ROCK)-mediated phosphorylation of myosinII regulatory light chain (RLC), RhoA-mediated its sustenance of RLC phosphorylation and activation of formin mDia are critical triggering events [17][18]. For more details, see Functional module: Integrin β1/syndecan-4 synergy in membrane protrusion and adhesion dynamics. Myosin IIA and Myosin IIB cooperatively aid actomyosin bundling in a localized manner thus stabilizing the large adhesions [19]. As a result, actin bundles assemble between stable adhesions on the ventral cell surface, forming structures known as stress fibers ([11], reviewed in [20]). Assembly of these stress fibers in the lamellum acts as a structural template for the compositional maturation of FAs and seems to be quintessential despite an intact actomyosin contractile mechanism [21]. At this stage, the adhesion components contain modifications that inhibit Rac-GEF recruitment leading to a local and hence define the cell rear [19]. Stress fibers participate in mechanotransduction by FAs [22] and can generate high level of forces on rigid and dense matrices [23][24], through actomyosin contractions.

Accumulation of tensional forces (internal or external) that results in FA reinforcement has been shown to ultimately activate and recruit Rho-GEFs to adhesion sites [25]. The two GEFs studied seem to be under the control of distinct pathways- Fyn (a SFK) and FAK-Ras-ERK signaling. Subsequent force-dependent Rho activation leads to events aiding the maturation process.

Thus, bidirectional force transmission between the internal cytoskeletal network and the exterior gets directed through the adhesions [26] (reviewed in [14][15]). Force differences across adhesions influence actin dynamics and signal transduction cascades, leading to the reinforcement and maturation of adhesions (reviewed in [22]).

During growth and early maturation, the FA size correlates with traction stress but can exert variable force depending on its position in the cell [27]. However, upon maturation, the changes in morphology, protein composition and/or phosphorylation of FAs decrease propulsive forces [7]. They do not grow further under tension [27] probably due to altered chemical state of FA signaling components [28].

Thus mature FAs effectively become passive anchorage devices for the maintenance of a spread cell morphology [7] and can sustain up to a six-fold increase in external force load without elongating [27]. Stability of receptor-stress fiber interactions enables mature adhesions to probe their environment over large spatial and temporal scales and to reorient, a feature essential for direction-dependent rigidity sensing [29][30].

FAs typically disassemble within 10 to 20 minutes or mature further into fibrillar adhesions [31][32] that can remodel the extracellular matrix. Alternately, high Src activity suppresses integrin function and contractility, thus fail to assemble the matrix and mediate transformation into podosomes [33].

References

1. Nobes CD, and Hall A. Rho, rac, and cdc42 GTPases regulate the assembly of multimolecular focal complexes associated with actin stress fibers, lamellipodia, and filopodia. Cell 1995; 81(1):53-62. [PMID: 7536630]
2. Clark EA, King WG, Brugge JS, Symons M, and Hynes RO. Integrin-mediated signals regulated by members of the rho family of GTPases. J. Cell Biol. 1998; 142(2):573-86. [PMID: 9679153]
3. Rottner K, Hall A, and Small JV. Interplay between Rac and Rho in the control of substrate contact dynamics. Curr. Biol. 1999; 9(12):640-8. [PMID: 10375527]
4. Zaidel-Bar R, Milo R, Kam Z, and Geiger B. A paxillin tyrosine phosphorylation switch regulates the assembly and form of cell-matrix adhesions. J. Cell. Sci. 2006; 120(Pt 1):137-48. [PMID: 17164291]
5. Ballestrem C, Erez N, Kirchner J, Kam Z, Bershadsky A, and Geiger B. Molecular mapping of tyrosine-phosphorylated proteins in focal adhesions using fluorescence resonance energy transfer. J. Cell. Sci. 2006; 119(Pt 5):866-75. [PMID: 16478788]
6. Giannone G, and Sheetz MP. Substrate rigidity and force define form through tyrosine phosphatase and kinase pathways. Trends Cell Biol. 2006; 16(4):213-23. [PMID: 16529933]
7. Beningo KA, Dembo M, Kaverina I, Small JV, and Wang YL. Nascent focal adhesions are responsible for the generation of strong propulsive forces in migrating fibroblasts. J. Cell Biol. 2001; 153(4):881-8. [PMID: 11352946]
8. Zaidel-Bar R, Ballestrem C, Kam Z, and Geiger B. Early molecular events in the assembly of matrix adhesions at the leading edge of migrating cells. J. Cell. Sci. 2003; 116(Pt 22):4605-13. [PMID: 14576354]
9. Ballestrem C, Hinz B, Imhof BA, and Wehrle-Haller B. Marching at the front and dragging behind: differential alphaVbeta3-integrin turnover regulates focal adhesion behavior. J. Cell Biol. 2001; 155(7):1319-32. [PMID: 11756480]
10. Choi CK, Vicente-Manzanares M, Zareno J, Whitmore LA, Mogilner A, and Horwitz AR. Actin and alpha-actinin orchestrate the assembly and maturation of nascent adhesions in a myosin II motor-independent manner. Nat. Cell Biol. 2008; 10(9):1039-50. [PMID: 19160484]
11. Chrzanowska-Wodnicka M, and Burridge K. Rho-stimulated contractility drives the formation of stress fibers and focal adhesions. J. Cell Biol. 1996; 133(6):1403-15. [PMID: 8682874]
12. Riveline D, Zamir E, Balaban NQ, Schwarz US, Ishizaki T, Narumiya S, Kam Z, Geiger B, and Bershadsky AD. Focal contacts as mechanosensors: externally applied local mechanical force induces growth of focal contacts by an mDia1-dependent and ROCK-independent mechanism. J. Cell Biol. 2001; 153(6):1175-86. [PMID: 11402062]
13. Casino FG, Gaudiano V, Santarsia G, Ambrosecchia R, Mostacci SD, Bagnato C, and Lopez T. Reconciliation between urea kinetics and direct dialysis quantification. Nephrol. Dial. Transplant. 1992; 7(10):1007-12. [PMID: 1331874]
14. Vicente-Manzanares M, Ma X, Adelstein RS, and Horwitz AR. Non-muscle myosin II takes centre stage in cell adhesion and migration. Nat. Rev. Mol. Cell Biol. 2009; 10(11):778-90. [PMID: 19851336]
15. Parsons JT, Horwitz AR, and Schwartz MA. Cell adhesion: integrating cytoskeletal dynamics and cellular tension. Nat. Rev. Mol. Cell Biol. 2010; 11(9):633-43. [PMID: 20729930]
16. Roca-Cusachs P, Iskratsch T, and Sheetz MP. Finding the weakest link: exploring integrin-mediated mechanical molecular pathways. J. Cell. Sci. 2012; 125(Pt 13):3025-38. [PMID: 22797926]
17. Narumiya S, Tanji M, and Ishizaki T. Rho signaling, ROCK and mDia1, in transformation, metastasis and invasion. Cancer Metastasis Rev. 2009; 28(1-2):65-76. [PMID: 19160018]
18. Beningo KA, Hamao K, Dembo M, Wang Y, and Hosoya H. Traction forces of fibroblasts are regulated by the Rho-dependent kinase but not by the myosin light chain kinase. Arch. Biochem. Biophys. 2006; 456(2):224-31. [PMID: 17094935]
19. Vicente-Manzanares M, Newell-Litwa K, Bachir AI, Whitmore LA, and Horwitz AR. Myosin IIA/IIB restrict adhesive and protrusive signaling to generate front-back polarity in migrating cells. J. Cell Biol. 2011; 193(2):381-96. [PMID: 21482721]
20. Le Clainche C, and Carlier M. Regulation of actin assembly associated with protrusion and adhesion in cell migration. Physiol. Rev. 2008; 88(2):489-513. [PMID: 18391171]
21. Oakes PW, Beckham Y, Stricker J, and Gardel ML. Tension is required but not sufficient for focal adhesion maturation without a stress fiber template. J. Cell Biol. 2012; 196(3):363-74. [PMID: 22291038]
22. Geiger B, Spatz JP, and Bershadsky AD. Environmental sensing through focal adhesions. Nat. Rev. Mol. Cell Biol. 2009; 10(1):21-33. [PMID: 19197329]
23. Rajagopalan P, Marganski WA, Brown XQ, and Wong JY. Direct comparison of the spread area, contractility, and migration of balb/c 3T3 fibroblasts adhered to fibronectin- and RGD-modified substrata. Biophys. J. 2004; 87(4):2818-27. [PMID: 15454473]
24. Yeung T, Georges PC, Flanagan LA, Marg B, Ortiz M, Funaki M, Zahir N, Ming W, Weaver V, and Janmey PA. Effects of substrate stiffness on cell morphology, cytoskeletal structure, and adhesion. Cell Motil. Cytoskeleton 2005; 60(1):24-34. [PMID: 15573414]
25. Guilluy C, Swaminathan V, Garcia-Mata R, O’Brien ET, Superfine R, and Burridge K. The Rho GEFs LARG and GEF-H1 regulate the mechanical response to force on integrins. Nat. Cell Biol. 2011; 13(6):722-7. [PMID: 21572419]
26. Wang N, and Suo Z. Long-distance propagation of forces in a cell. Biochem. Biophys. Res. Commun. 2005; 328(4):1133-8. [PMID: 15707995]
27. Stricker J, Aratyn-Schaus Y, Oakes PW, and Gardel ML. Spatiotemporal constraints on the force-dependent growth of focal adhesions. Biophys. J. 2011; 100(12):2883-93. [PMID: 21689521]
28. Prager-Khoutorsky M, Lichtenstein A, Krishnan R, Rajendran K, Mayo A, Kam Z, Geiger B, and Bershadsky AD. Fibroblast polarization is a matrix-rigidity-dependent process controlled by focal adhesion mechanosensing. Nat. Cell Biol. 2011; 13(12):1457-65. [PMID: 22081092]
29. Walcott S, and Sun SX. A mechanical model of actin stress fiber formation and substrate elasticity sensing in adherent cells. Proc. Natl. Acad. Sci. U.S.A. 2010; 107(17):7757-62. [PMID: 20385838]
30. Zemel A, Rehfeldt F, Brown AEX, Discher DE, and Safran SA. Optimal matrix rigidity for stress fiber polarization in stem cells. Nat Phys 2010; 6(6):468-473. [PMID: 20563235]
31. Laukaitis CM, Webb DJ, Donais K, and Horwitz AF. Differential dynamics of alpha 5 integrin, paxillin, and alpha-actinin during formation and disassembly of adhesions in migrating cells. J. Cell Biol. 2001; 153(7):1427-40. [PMID: 11425873]
32. Webb DJ, Donais K, Whitmore LA, Thomas SM, Turner CE, Parsons JT, and Horwitz AF. FAK-Src signalling through paxillin, ERK and MLCK regulates adhesion disassembly. Nat. Cell Biol. 2004; 6(2):154-61. [PMID: 14743221]
33. Huveneers S, Arslan S, van de Water B, Sonnenberg A, and Danen EHJ. Integrins uncouple Src-induced morphological and oncogenic transformation. J. Biol. Chem. 2008; 283(19):13243-51. [PMID: 18326486]